PT - JOURNAL ARTICLE AU - Priyavathi Dhandapani AU - Srujan Kumar Dondapati AU - Anne Zemella AU - Dennis Bräuer AU - Doreen Anja Wüstenhagen AU - Stefan Kubick TI - Targeted Esterase induced Dye loading supports Calcium Imaging in Eukaryotic Cell-Free Systems AID - 10.1101/2020.03.05.978247 DP - 2020 Jan 01 TA - bioRxiv PG - 2020.03.05.978247 4099 - http://biorxiv.org/content/early/2020/03/06/2020.03.05.978247.short 4100 - http://biorxiv.org/content/early/2020/03/06/2020.03.05.978247.full AB - Calcium imaging is an important functional tool for addressing ion channels, transporters and pumps for drug screening in living cells. Depicted eukaryotic cell-free systems utilize microsomes, derived from endoplasmic reticulum to incorporate the synthesized membrane proteins. Absence or inadequate amount of carboxylesterase in the endoplasmic reticulum of eukaryotic cells, which is necessary to cleave the acetoxymethyl ester moiety of the chemical calcium indicators, advocates the hindrance to perform calcium imaging in microsomes. In this work, we try to overcome this drawback and adapt the cell-based calcium imaging principle to a cell-free protein synthesis platform. Carboxylesterase synthesized in a Spodoptera frugiperda Sf21 lysate translation system is established as a viable calcium imaging tool and hTRPV1 is used as a model channel protein to demonstrate the realization of this concept.